Background/Purpose: In osteoarthritis (OA), defects in Autophagy are evident and precede joint damage. Therefore, identifying hallmarks associated with specific autophagy subtypes could shed light to fundamental mechanisms of joint disease.

Methods: A comparative analysis of 35 autophagy genes was performed from blood from the Prospective OA Cohort of A Coruña (PROCOAC). Non-OA subjects (n=18) and Knee OA subjects (n=18, OA III-IV) were profiled using an autophagy gene expression array. Confirmatory studies were performed in blood from Non-OA subjects (n=30) and Knee-OA subjects (n=30, OA III-IV).

The candidate gene was evaluated in human knee joint tissues (cartilage, meniscus, ligaments, synovium) with different KL grades (KL0, 2, 4) and in both spontaneous aging and surgically-induced OA in mice by IHC. The functional consequences were studied in primary human OA chondrocytes. Autophagy, Chaperone-mediated autophagy (CMA), inflammation, and cellular senescence were analyzing by gene and protein expression. Moreover, oxidative stress and cell death were evaluated by FACS. The contribution of CMA to chondrocyte homeostasis was evaluated by studying the capacity of CMA to restore proteostasis upon autophagy deficiency by siATG5.

Results: 15 autophagy-related genes were significantly downregulated in blood from knee OA patients compared to non-OA patients. No significant upregulation was found for any studied gene, although a trend towards upregulation was found in genes involved in the mTOR pathway. Interestingly, HSP90AA1 and HSPA8, CMA markers involved in stress response and protein folding, were downregulated. Confirmatory studies showed a significant downregulation of MAP1LC3B and HSP90AA1 in blood from knee OA patients. Remarkably, HSP90A was found reduced in femoral cartilage, meniscus and ACL, while in synovial membrane, the expression was found increased. This expression signature was dependent on OA severity. In addition, we observed a decrease of HSP90A with aging and OA in mice. The functional consequences of HSP90AA1 gene silencing are related to an increase in NFkB, MMP13, and p16 expression. Interestingly, LAMP2A, a key CMA marker, and MAP1LC3B expression were upregulated, which might indicate an early response to maintain homeostasis. On the other hand, LAMP2A protein is decreased upon HSP90AA1 deficiency, while LC3II and p62 were increased, indicating a failure in the autophagy flux that leads to impaired lysosomal degradation. Moreover, p21, p16 and prbS6 were increased, besides increasing mitochondrial ROS production and apoptosis. ATG5 silencing blocks autophagy by reducing LC3II and increasing prbs6, p62, p16 and p21. Interestingly, LAMP2A and HSP90A were found increased, indicating a compensative activation of CMA in response to autophagy defects. These results support that HSP90A has an important role in chondrocyte homeostasis by participating in the cross-talk between CMA and autophagy.

Conclusion: Taking together, we identified HSP90A, a CMA regulator, as critical in chondrocyte homeostasis. These disease mechanisms are relevant in OA and constitute hallmarks potentially useful to prevent OA progression.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/chaperone-mediated-autophagy-is-a-hallmark-of-joint-damage-in-osteoarthritic-patients/